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NMR based CSF metabolomics in tuberculous meningitis: correlation with clinical and MRI findings

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Abstract

We report the potential role of 1H Nuclear Magnetic Resonance (NMR) based metabolomics in tuberculous meningitis (TBM). We also correlate the significant metabolites with clinical-radiological parameters. Forty-three patients with TBM were included, and their severity of meningitis was graded as stages I to III, and patients with positive Mycobacterium tuberculosis or its nucleic acid was considered as definite TBM. 1H NMR-based metabolomic study was performed on (CSF) samples, and the significant metabolites compared to healthy controls were identified. Outcome at three months was defined as death, poor and good based on the modified Rankin Scale. These metabolites were compared between definite and probable groups of TBM, and also correlated with MRI findings. About 11 metabolites were found to be significant for distinguishing TBM from the controls. In TBM, lactate, glutamate, alanine, arginine, 2-hydroxyisobutyrate, formate, and cis-aconitate were upregulated, and glucose, fructose, glutamine, and myo-inositol were downregulated compared to the controls. For differentiating TBM from the controls, the AUC of the ROC curve generated using these significant metabolites was 0.99, with a 95% confidence interval from 0.96 to 1, demonstrating that these metabolites were able to classify cases with good sensitivity and specificity. Lactate concentration in CSF correlated with hemoglobin, CSF glucose, and infarction. The outcome did not correlate with metabolomics parameters. NMR-based CSF metabolomics have a potential role in differentiating TBM from the controls.

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Acknowledgments

We thank Mr. Shakti Kumar for his secretarial help.

Funding support

This work was supported by an Intramural Grant (A-04-PGI/IMP/76/2018) of Sanjay Gandhi Postgraduate Institute of Medical Sciences, Lucknow, Sanctioned to Professor Jayantee Kalita.

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Authors and Affiliations

  1. Department of Neurology, Sanjay Gandhi Postgraduate Institute of Medical Sciences, Lucknow, Uttar Pradesh, 226014, India

    Ruchi Shukla, Jayantee Kalita & Usha Kant Misra

  2. Centre of Biomedical Research, Lucknow, Uttar Pradesh, 226014, India

    Rashmi Parihar & Bikash Baishya

  3. Department of Anaesthesiology, Sanjay Gandhi Postgraduate Institute of Medical Sciences, Lucknow, India

    Rudrashish Haldar

  4. Department of Bioinformatics, Dr. A. P. J. Abdul Kalam Technical University, Lucknow, India

    Rashmi Parihar

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  1. Rashmi Parihar
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Contributions

RP: NMR experiment and data analysis, RS: Data collection and analysis of data, BB: Supervising the experimental data and their result, and writing the manuscript. JK: management, clinical data collection, interpretation of data, writing and drafting the manuscript. RH: Provided control CSF sample. UKM: Supervision, patient management, and writing the manuscript.

Corresponding authors

Correspondence to Bikash Baishya or Jayantee Kalita.

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Ethical approval

This study was ethically approved by the institutional ethics committee, SGPGIMS, Lucknow, India (2018–28-IMP-102, dated 13-4-2018).

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Supplementary Information

Supplementary Fig. 1.

A) PCA 2D score plot of TBM (Definite vs. Probable) displaying significant overlap as both are disease subgroup, B) PLS-DA score plot of TBM (Definite vs. Probable), C) VIP plot of TBM (Definite vs. Probable, D) PLS-DA loading plot of TBM (Definite vs. Probable). Lactate and glucose are the most important metabolites for discrimination, followed by glutamate, glutamine, o-phosphoserine, arginine, and malonate. (TIFF 254 kb) (PNG 6534 kb)

High resolution image (TIFF 6438 kb)

Supplementary Fig. 2.

A) PCA 2D score plot showing A) control vs. TBM, B) control Vs. definite, and C) Control vs. probable. There is clear clustering between the two groups; the best separation is observed between control vs. definite TBM. (PNG 14202 kb)

High resolution image (TIFF 5006 kb)

Supplementary Fig. 3.

Stack plot of a series of 1H1D spectra(cpmg). Panels a-i are expanded sections from control CSF, and panels j-s are from definite TBM cases. Similar observations in dd at 4.45 is shown with dotted box were also observed. The identity of this spin system could not be confirmed from the databases. In one case panel, 'r' a quartet at 4.43 was observed, which may be from acetoin- an external energy store for bacteria. This peak could not be detected in other samples. (PNG 2991 kb)

High resolution image (TIFF 593 kb)

Supplementary Fig. 4.

Panels A-F has expended portions of the various regions of the tilted J-RES spectrum. Numbers near each peak position represent metabolites from the metabolites list in supplementary table 1. (PNG 2751 kb)

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Supplementary Fig. 5.

An expended portion of the 1H-13C HSQC spectrum showing all peaks that were assigned. Individual peak assignments are further displayed in supplementary Figs 7-10. (PNG 991 kb)

High resolution image (TIFF 236 kb)

Supplementary Fig. 6.

(A) Full 1H-1H TOCSY spectrum, (B) Expended portion of the 1H-1H TOCSY spectrum showing the important region that was assigned. Individual peak assignments are further displayed in supplementary Figs 11-13. (C) Expanded portion showing the cross-peaks between the peak at 4.45 with 1.75 and 0.88 ppm. Half of the TBM cases displayed this peak at 4.45, while no such peak was observed in the control group. (PNG 1426 kb)

High resolution image (TIFF 254 kb)

Supplementary Fig. 7.

Expended portion for each peak from 1H-13C HSQC spectrum that was assigned. Numbers represent metabolites from supplementary table 1. (PNG 1028 kb)

High resolution image (TIFF 232 kb)

Supplementary Fig. 8.

Expended portion for each peak from 1H-13C HSQC spectrum that was assigned. Numbers represent metabolites from supplementary table 1. (PNG 1078 kb)

High resolution image (TIFF 204 kb)

Supplementary Fig. 9.

Expended portion for each peak from 1H-13C HSQC spectrum that was assigned. Numbers represent metabolites from supplementary table 1. (PNG 1153 kb)

High resolution image (TIFF 233 kb)

Supplementary Fig. 10.

Expended portion for each peak from 1H-13C HSQC spectrum that was assigned. Numbers represent metabolites from supplementary table 1. (PNG 950 kb)

High resolution image (TIFF 189 kb)

Supplementary Fig. 11.

Expended portion for each peak from 1H-1H TOCSY spectrum that was assigned. Numbers represent metabolites from supplementary table 1. (PNG 1174 kb)

High resolution image (TIFF 237 kb)

Supplementary Fig. 12.

Expended portion for each peak from 1H-1H TOCSY spectrum that was assigned. Numbers represent metabolites from supplementary table 1. (PNG 784 kb)

High resolution image (TIFF 143 kb)

Supplementary Fig. 13.

Expended portion for each peak from 1H-1H TOCSY spectrum that was assigned. Numbers represent metabolites from supplementary table 1. (PNG 1206 kb)

High resolution image (TIFF 246 kb)

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Parihar, R., Shukla, R., Baishya, B. et al. NMR based CSF metabolomics in tuberculous meningitis: correlation with clinical and MRI findings. Metab Brain Dis 37, 773–785 (2022). https://doi.org/10.1007/s11011-021-00860-y

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